Domestic animal hosts strongly influence human-feeding rates of the chagas disease vector Triatoma...

Domestic Animal Hosts Strongly Influence Human-Feeding Rates of the Chagas Disease Vector Triatomainfestans in ArgentinaRicardo E. Gurtler1*, Marıa C. Cecere1, Gonzalo M. Vazquez-Prokopec1,2, Leonardo A. Ceballos1,

Juan M. Gurevitz1, Marıa del Pilar Fernandez1, Uriel Kitron2, Joel E. Cohen3

1 Laboratory of Eco-Epidemiology, Department of Ecology, Genetics and Evolution, Universidad de Buenos Aires-IEGEBA (CONICET-UBA), Buenos Aires, Argentina,

2 Department of Environmental Studies, Emory University, Atlanta, Georgia, United States of America, 3 Laboratory of Populations, Rockefeller and Columbia Universities,

New York, New York, United States of America

Abstract

Background: The host species composition in a household and their relative availability affect the host-feeding choices ofblood-sucking insects and parasite transmission risks. We investigated four hypotheses regarding factors that affect blood-feeding rates, proportion of human-fed bugs (human blood index), and daily human-feeding rates of Triatoma infestans, themain vector of Chagas disease.

Methods: A cross-sectional survey collected triatomines in human sleeping quarters (domiciles) of 49 of 270 rural houses innorthwestern Argentina. We developed an improved way of estimating the human-feeding rate of domestic T. infestanspopulations. We fitted generalized linear mixed-effects models to a global model with six explanatory variables (chickenblood index, dog blood index, bug stage, numbers of human residents, bug abundance, and maximum temperature duringthe night preceding bug catch) and three response variables (daily blood-feeding rate, human blood index, and dailyhuman-feeding rate). Coefficients were estimated via multimodel inference with model averaging.

Findings: Median blood-feeding intervals per late-stage bug were 4.1 days, with large variations among households. Themain bloodmeal sources were humans (68%), chickens (22%), and dogs (9%). Blood-feeding rates decreased with increasesin the chicken blood index. Both the human blood index and daily human-feeding rate decreased substantially withincreasing proportions of chicken- or dog-fed bugs, or the presence of chickens indoors. Improved calculations estimatedthe mean daily human-feeding rate per late-stage bug at 0.231 (95% confidence interval, 0.157–0.305).

Conclusions and Significance: Based on the changing availability of chickens in domiciles during spring-summer and themuch larger infectivity of dogs compared with humans, we infer that the net effects of chickens in the presence oftransmission-competent hosts may be more adequately described by zoopotentiation than by zooprophylaxis. Domesticanimals in domiciles profoundly affect the host-feeding choices, human-vector contact rates and parasite transmissionpredicted by a model based on these estimates.

Citation: Gurtler RE, Cecere MC, Vazquez-Prokopec GM, Ceballos LA, Gurevitz JM, et al. (2014) Domestic Animal Hosts Strongly Influence Human-Feeding Rates ofthe Chagas Disease Vector Triatoma infestans in Argentina. PLoS Negl Trop Dis 8(5): e2894. doi:10.1371/journal.pntd.0002894

Editor: Eric Dumonteil, Universidad Autonoma de Yucatan, Mexico

Received August 7, 2013; Accepted April 14, 2014; Published May 22, 2014

Copyright: � 2014 Gurtler et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permitsunrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

Funding: This study was supported by awards from the Agencia Nacional de Promocion Cientıfica y Tecnica of Argentina (PICTO-Glaxo 2011-0062 and PICT-2011-2072) to REG; National Institutes of Health/National Science Foundation Ecology of Infectious Disease program award R01 TW05836 funded by the FogartyInternational Center and the National Institute of Environmental Health Sciences to UK, REG, and JEC; by the University of Buenos Aires (REG). The funders had norole in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

Competing Interests: The authors have declared that no competing interests exist.

* E-mail: [email protected]

Introduction

The host species composition in a habitat, relative host abundance

and proximity affect the host-feeding choices of hematophagous insect

vectors and associated parasite transmission risks [1]. Host defensive

behavior in response to attacks and the density of blood-sucking

insects also modify host-feeding patterns in triatomine bugs, vector of

Chagas disease [2], and in blackflies, vectors of river blindness [3].

The estimation of host choices and blood-feeding rates, two key

parameters used to estimate vectorial capacity in mathematical

models of vector-borne diseases [4], is fraught with difficulties [5].

Zooprophylaxis is the use of wild or domestic animals, which

are not the reservoir hosts of a given disease, to divert the blood-

seeking mosquito vectors from the human hosts of that disease [6].

Its use for malaria prevention dates back to the early twentieth

century [7], and was strongly advocated as a supplementary

control strategy in the 1980s [8]. Empirical evidence showed

conflicting results in various parts of the world. Cattle sometimes

increased, and sometimes decreased, the proportion of Anopheles

mosquitoes that fed on humans [9,10 and references therein].

Although the malaria vector Anopheles arabiensis took a high

proportion of meals from cattle, the vector population fed

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sufficiently on humans to transmit malaria and therefore classical

zooprophylaxis may not have a significant impact [11]. Mathe-

matical models predicted that under certain conditions of strong

host-feeding preference for the protective host, the diversion of

mosquito vectors would reduce malaria transmission [12].

Computer simulations predicted that reducing the mosquitoes’

access to humans would have a much greater effect on the

transmission of malaria than for Japanese encephalitis (which has

animal reservoir hosts), where the most critical factor would be the

proximity of the reservoir hosts to vector breeding sites [13]. For

sand fly vectors of visceral leishmaniasis, the effects of cattle on

vector abundance and transmission were also mixed [14]. For tick-

borne Lyme disease in northeastern USA, the relative proportion

of white-footed mice and Virginia opossums was predicted to

affect transmission risks [15], but the net outcome may depend on

the presence of other more or less competent hosts [7].

Both the concept of zooprophylaxis and its more recent relative,

the "dilution effect" [16], propose that the probability of vectors

feeding on transmission-competent hosts would decline with

increasing numbers of non-competent hosts. Non-competent hosts

would also reduce the abundance of infected vectors and

transmission risks under certain, not all, circumstances [7,17].

The opposite of zooprophylaxis is zoopotentiation [13]: exacer-

bated parasite transmission by wild or domestic animals that are

not the reservoir hosts of a given disease.

Triatoma infestans –the most important vector of the kinetoplastid

protozoan Trypanosoma cruzi that causes Chagas disease in humans

and mammalian hosts– is well adapted to thrive in human sleeping

quarters and surrounding habitats occupied by domestic animals

[18,19]. This trait, combined with epidemiological significance,

susceptibility to modern pyrethroids and political momentum,

justified targeting T. infestans for elimination in the southern cone

countries of South America since 1991 [19]. Large-scale insecti-

cide spraying campaigns suppressed this species from extensive

areas in Brazil, Chile and Uruguay where transmission of human

T. cruzi infection through T. infestans was interrupted [20].

Elsewhere in the southern cone countries and more particularly

in the Gran Chaco ecoregion extending over Argentina, Bolivia

and Paraguay, insecticide control campaigns have reduced but not

eliminated transmission of human T. cruzi infection [21]. Although

the available information is sparse and disease control status

heterogeneous, the magnitude of the problem is reflected in the

high seroprevalence of T. cruzi in pregnant women examined

mostly at public health services from 1999 to 2004, which ranged

from 5 to 20% in the most affected provinces in the Argentinean

Chaco [22]. Regarding vector-borne transmission, annual inci-

dence rates of human infection rose to 4.3-4.6% in some highly

infested regions of the Argentinean and Bolivian Chaco [23,24].

The notion of zooprophylaxis has been debated in Chagas

disease [2,25 and references therein]. In human sleeping quarters,

T. infestans mainly feeds on humans, dogs, chickens and cats

throughout its distribution range [2,26,27]. Chickens and other

birds are not susceptible to T. cruzi and avian blood exerts no

effects on an established bug infection (though it supports the

development and reproduction of bugs), whereas dogs and cats are

key reservoir hosts of T. cruzi [21,25]. In households from three

rural villages in northwestern Argentina, the proportion of

domestic T. infestans that blood-fed on humans (i.e., human blood

index) decreased substantially with the increasing presence of

chickens or dogs in human sleeping quarters and with increasing

domestic bug abundance during spring-summer [2]. ‘‘Domestic

bug’’ refers to bugs captured in the set of contiguous human

sleeping quarters and rooms that share a continuous roof structure

(i.e., domestic areas or domiciles). The presence of chickens

nesting in domestic areas correlated positively with increased bug

abundance and with an increasing proportion of chicken-fed bugs

[28]; reduced the proportion of T. cruzi-infected domestic T.

infestans, and increased the abundance of infected vectors [25].

Moreover, the adjusted odds of human infection with T. cruzi

increased with the household number of infected dogs or cats and

the presence of chickens indoors [29]. An experimental study

corroborated that the additional presence of chickens relative to

infected guinea pigs reduced bug infection prevalence [30]. A

mathematical model of the household transmission of T. cruzi was

consistent with this empirical evidence and predicted that the

fraction of spring bugs’ feeding contacts with humans would

decrease with more dogs and chickens in spring because both

animal hosts would divert vector feeding contacts [31]. This

prediction remained untested in the absence of an appropriate

method to estimate blood-feeding rates and field data.

There is a striking lack of information on the blood-feeding rates

of Triatominae in field conditions. Feeding rates of domestic T.

infestans and Rhodnius prolixus were first estimated indirectly by the

distributions of bug weight and length in 2 and 14 houses,

respectively [32,33]. A more widely applicable method estimated

daily feeding rates by measuring the temperature-adjusted

occurrence of transparent (clear) urine assessed shortly after

capture [34]. Using this method, domestic populations of T.

infestans were estimated to blood-feed every three days over the

spring-summer period [35], and those from chicken coops fed

more often than bugs from other peridomestic habitats in spring

[36,37]. The product of the human blood index and daily feeding

rate is not a daily rate because of the undefined period of time

during which the bugs may have fed on a given host species, and

because the human blood index cannot distinguish whether the

bugs fed once or several times on humans [24]. Therefore, in this

study we developed a different way of calculating human-feeding

rates that provides more informative estimates and a measure of its

variability among houses in a population-based survey.

We also collected additional data to estimate the daily blood-

feeding rates, host choices, and rates of human-bug feeding

contacts per person of domestic T. infestans from the same study

Author Summary

The major vectors of Chagas disease are species oftriatomine bugs that have adapted to human sleepingquarters and may feed on domestic animals and humans.There is a striking lack of information on the blood-feedingrates of Triatominae in field conditions, and factorsmodifying the fraction of bugs that feed on humans haverarely been investigated. Here we tested whether thespring fraction of bugs’ feeding contacts with humanswould decrease when dogs and chickens are available inhuman sleeping quarters in a well-defined endemic ruralarea in northwestern Argentina. On average, late-stagebugs fed every four days in spring and the majority fed onhumans. The results demonstrate that both the percent-age of human-fed bugs and the daily rate of feeding onhumans decreased substantially with increasing propor-tions of chicken- or dog-fed bugs in the house, or thepresence of chickens indoors. Combined with earlierfindings on the seasonal dynamics of bug abundance,host-feeding patterns and the relative infectivity ofdomestic hosts, the net effects of the presence of chickensand dogs in human sleeping quarters is predicted toincrease the transmission of T. cruzi in summer whenchickens move outdoors.

Human-Feeding Rates of Triatoma infestans


region as our previous investigations, although in the context of

lower domestic infestations. Factors modifying the fraction of bugs

that feed on humans have rarely been investigated. Using

generalized linear mixed-effects models (GLMM) and a multi-

model inference approach with model averaging, here we test the

following hypotheses related to domestic T. infestans: 1) Blood-

feeding rates should increase with host availability (i.e., humans,

chickens, dogs) and ambient temperature [38]; 2) The human

blood index and human-feeding rate should decrease with more

chicken or dog blood meals and the occurrence of indoor-nesting

chickens and dogs (i.e., greater host diversity and abundance) [2];

3) The human blood index and human-feeding rate should

decrease with increasing bug abundance per unit of sampling

effort because of increasing host irritation with increasing bites per

host [2], and 4) The predicted number of human-bug feeding

contacts per person-day should decrease with the indoor presence

of chickens and dogs in spring [31]. Testing these hypotheses is

important for risk assessment and a better understanding and

control of domestic populations of Triatominae. Our current

results support and extend previous findings and the main

predictions of the mathematical model.

Materials and Methods

Study areaField work was carried out in October-November 2003 in eight

neighboring rural communities with 270 houses in Figueroa

Department (27u 239S, 63u 299W), Province of Santiago del

Estero, Argentina, described elsewhere [39]. The study area was

selected for an insecticide trial in 2003–2005 because it had high

infestation with T. infestans and recent occurrence of human acute

cases of T. cruzi despite a previous community-wide residual

spraying with pyrethroid insecticides conducted in 2000. Most

houses were made of adobe walls and thatched roofs, with one or

two adjacent bedrooms and a front verandah 5–10 m wide, and

had multiple peridomestic structures for housing domestic animals.

A weather station (Weather Monitor II, Davis, Baltimore, MD)

located 50 km from the study area measured temperature, relative

humidity, wind speed and direction, barometric pressure, and

rainfall at 15 min intervals.

Study designA cross-sectional survey of house infestation was conducted

before control interventions in October-November 2003 [39].

Each of the 270 houses was visited and georeferenced. The

location and type of building material of each structure were

recorded in a sketch map and in a form. All houses positive for T.

infestans during the first week of the survey (October 20–27, 2003)

were considered eligible for blood-feeding studies. Time con-

straints for processing the bugs within 8 hours of capture (needed

to estimate blood-feeding frequency) dictated that insects from

domestic areas at 49 different house compounds were processed

(i.e., 52% of houses with domestic areas found infested). These 49

sites were all of the sites that satisfied the time constraints, and

were not a sample of a larger number of possible sites.

Householders were asked about the number of human

occupants aged 15 years or more and less than 15, and of the

domestic animals they owned, their resting places at night, and

whether chickens, dogs and cats were allowed indoors or in the

verandah. We requested information on chickens’ current and

previous nesting sites and sought direct evidence on their current

nesting and resting sites.

Four teams, each one composed of one member of the research

team and three skilled bug collectors, searched for triatomine bugs

in all sites using timed manual collections with a dislodging spray

(0.2% tetramethrin, Espacial 0.2, Buenos Aires). One person

searched systematically in human sleeping quarters during 30 min

(0.5 person-hour per domicile) whereas two persons searched

peridomestic structures. Searches were usually conducted between

8:00 and 14:00 hours on non-rainy days, but sometimes they had

to start later because householders were not available.

The weather station measured the maximum temperature

during each 15-minute interval. It then reported the two-hour

mean maximum temperature as the average of these eight

measurements. We calculated the mean maximum temperature

from 20:00 to 6:00 h as the grand mean of the mean maxima

recorded in each two-hour period (data in Table S1). Our mean

maximum temperatures averaged 25.4uC (SD, 3.0; range of two-

hour period means, 21.9–30.0uC) and wind speed averaged

3.8 km/h (SD, 2.1; range of two-hour period means, 1.6–7.8 km/

h). At 20:00 h (when bug flight and host-seeking activity usually

starts or peaks), maximum temperatures averaged 31.9uC (SD,

4.9; range, 26.8–39.8uC) and wind speed averaged 5.2 km/h (SD,

3.4; range, 1.6–11.3 km/h). Therefore, weather conditions during

the period from 20:00 to 6:00 h preceding every bug collection

day were suitable for blood-feeding.

All triatomine bugs collected were stored in labeled plastic bags

and kept in a cooler at 10–12uC between capture and arrival at

the field laboratory to stop excretion, and then were identified to

species and counted by stage [39]. Timing of bug catches and

time of processing of the bugs from each study house were

recorded. Late stages (fourth- and fifth-instars and adults) were

individually examined for the presence of colorless (transparent or

clear) urine within 8 hours of capture using the method

developed by Catala [34]. The abdomen of each individual bug

was compressed with tweezers on a clean slide until obtaining a

drop of feces whose color was recorded. We excluded first- to

third-instar nymphs from the analyses because there was no

information on how long transparent urine was detectable after

blood-feeding.

To estimate daily blood-feeding rates, the house-specific

proportion of T. infestans that fed during the preceding night was

estimated as a weighted average of the observed proportion of

fourth- and fifth-instar nymphs with colorless urine multiplied by a

temperature-dependent correction factor c = 0.0533 * t–0.585 and

the (uncorrected) proportion of adult bugs with colorless urine

[34]. Temperatures (tuC) were calculated from the mean of

maxima recorded every two hours between 20:00 h of the day

preceding capture to the catch time. The temperature-dependent

factor c allowed for extended detection of transparent urine in

nymphs over two nights or so within the temperature range 18–

28uC. In adult bugs, the transparent urine did not last from one

day to the next. Specifically, if we collected and examined x fourth-

or fifth-instar nymphs, of which a proportion f had transparent

urine, and y adult bugs, of which a proportion g had transparent

urine, then the temperature-adjusted overall proportion of late-

stage bugs with transparent urine in this site was estimated as

h = (x*c*f+y*g)/(x+y). The feeding interval (in days) was calculated

as the reciprocal 1/h, assuming independence in feeding among

bugs, a constant probability of feeding on any given day, and

recently-fed bugs equally likely to be caught as not recently-fed

bugs. Then a weighted average of h was estimated over all sites

which had at least six insects examined for urine transparency. We

estimated median feeding intervals because a few sites had no bugs

with transparent urine, and therefore 1/h was indefinite; in these

cases feeding intervals were arbitrarily taken to be very long (30

days) and medians calculated. All bugs were kept frozen at 220uCupon arrival to the laboratory in Buenos Aires.



Identification of blood mealsThe individual blood meals of all late-stage bugs collected at

each house were prepared for testing by cutting the thorax

transversally at the level of the third pair of legs and then

extracting the midgut with the blood meal into a previously

labeled, weighed vial [40]. Our previous studies based on some

2,000 identified blood meals of domestic T. infestans found minor

differences among nymphal instars and adult stages [26]. Bug

stages were verified on dissection and errors detected in field data

sheets corrected. A direct ELISA assay was used to test bloodmeal

contents against human, dog, cat, chicken, pig, goat and murid

rodent (rat or mouse) antisera as described before [40]. Each

microtiter plate contained test and control samples of the target

host species and four negative controls (i.e., bloodmeal contents or

serum of heterologous host species in PBS). Samples were tested in

duplicates, and the outcome was considered valid if the mean of

both duplicates of each sample did not differ by more than 15%.

For each host-ELISA system, dilutions of heterologous sera were

added to the antibody-enzyme conjugate solutions to decrease

cross-reactivity as described in [40]. To describe how bugs fed on

the available hosts, we report the proportion of reactive bugs (i.e.,

those positive against any of the tested antisera) that contained

each type of host blood.

Sensitivity and specificity of the direct ELISA for the target host

species was evaluated by testing three batches of 44 blood meals

each from bugs fed to repletion separately on a dog, cat and

chicken, kept at room temperature, killed 7–30 days after feeding

and held frozen at 220uC; sensitivity was 100% and specificity

97–100% [40]. Additionally, we tested 5–10 sera from each of the

seven target host species using the same procedures; sensitivity and

specificity were 100% for all hosts. Maximum homologous serum

titers at which a positive result was obtained (i.e., sensitivity limits)

were higher than 200,000 for all the antisera.

Data management and analysisThe daily rate of feeding on humans only per late-stage

domestic bug was calculated as the proportion of human-fed bugs

with unmixed blood meals among all domestic bugs examined for

transparent urine, any bloodmeal content and bloodmeal source.

Bugs were coded as 1 if they had an unmixed human blood meal

and a recent blood-feed, and 0 otherwise (i.e., any bug that did not

have transparent urine or a human blood meal or which was

empty on dissection automatically scored as 0). Three bugs with

transparent urine and mixed blood meals including humans were

excluded because it could not be established whether the most

recent blood meal was on humans or not; 319 bugs (from 45

houses) had valid data for human-feeding rates. The daily human-

feeding rate per domestic late-stage bug divided by the total

number of resident humans at each household yielded the average

daily human-feeding rate per person per late-stage domestic bug.

To estimate the average number of human-bug feeding contacts

per person-day in each study household, we multiplied the average

human-feeding rate (per person-day per late-stage domestic bug)

with the abundance of late-stage bugs (per unit of catch effort)

divided by bug capture efficiency. We assumed tentatively that bug

capture efficiency is the same, whether or not a bug fed on people,

and that K person-hour would catch 10% of late stages, because

one person-hour of catch effort by qualified bug collectors using a

dislodging spray captured 10% of the total domestic population of

T. infestans (capture efficiency) as determined by subsequent partial

house demolition [41, based on unpublished results in the same

province under similar weather conditions].

The binomial response variables analyzed were: i) daily blood-

feeding rate (measuring the occurrence of transparent urine

relative to the number of bugs examined for urine color, adjusted

for temperature-dependent detection of transparent urine in late-

stage nymphs); ii) human blood index (i.e., the proportion of

reactive bugs with human blood meals detected, regardless of

other blood sources), and iii) daily human-feeding rate per person

per bug (as defined above). All proportions reported have attached

standard errors clustered by household as estimated by Stata 12

[42].

We used a multimodel approach with model averaging run in R

(version 2.15.2) [43] following the strategy outlined by Burnham &

Anderson [44] and methods detailed by Grueber et al. [45] with

packages lme4 (version 0.999999-2), MuMIn (version 1.9.5), arm

(version 1.6–06.01), ResourceSelection (0.2–2), and ROCR

(version 1.0–5). For this purpose we fitted GLMMs to a global

model that included all the explanatory variables. The random-

intercept regression models clustered by bug collection site

addressed the fact that insects from a given site shared the same

environment and other undetermined characteristics that may

have created dependencies among responses within the same

cluster of observations. All predictors were established a priori

based on existing empirical evidence reviewed elsewhere [2,40].

The global model fitted to the response variables (pij) was:

logit (pij) = ln (pij/(1-pij)) = a+b16chickbij+b26dogbij+b36hum-

totj+b46stage3ij+b56maxtempj+b66bugabundj+aj,

where a is a constant; b1-b6 are regression coefficients; subscripts

are for insect i on house j; chickbi is the chicken blood index for

house j (a proportion); dogbi, the dog blood index for house j (a

proportion); humtot, the total number of human residents per

house; stage3, bug stage (with three levels: fourth- or fifth-instar

nymphs; adult males, and adult females); maxtemp, mean

maximum temperatures during the night preceding bug catch;

bugabund, domestic bug abundance per unit of catch effort

(including any first-third instar nymphs caught); aj, the random

intercept term, assumed to be independent across houses and

normally distributed with mean 0 and variance sa2 [46].

Predictors were not transformed for analysis.

We investigated potential multicollinearity problems of the

standardized variables (except stage) via variable inflation factors

(VIF), R2 and the condition number. For every predictor, VIFs

were ,1.25, R2,0.2, and the matrix had a condition number of

3.2, indicating no significant multicollinearity. For additional

analyses, we added the square of bug abundance to the global

model to test for non-linear effects; marginally significant effects

were detected only for daily feeding rates (results not shown). We

also replaced the chickbi and dogbi with the reported or observed

occurrence of chickens indoors and the number of dogs sharing

human sleeping quarters (‘‘roommate’’ dog) per household,

respectively, while keeping all other predictors the same.

For model selection we used Akaike’s Information Criterion

corrected for small samples (AICc). The subset of models that were

within 4 AICc from the best-fitting model were considered the top

models. The relative importance (RI) of each explanatory variable

in the top model set was computed using the Akaike weight for

each model in which the variable was present. The overall quality

of the fitted logistic regression models was assessed by means of the

Hosmer and Lemeshow test using the averaged coefficients, and

by the area under the receiver operating characteristic curve

(ROC). The latter can be interpreted as the fraction of all

observations that are correctly classified by the model.

We compared the outcome from the multimodel framework

with the more traditional approach based on null-hypothesis

testing via random-intercept logistic regression models clustered by

bug collection site as implemented in the command xtlogit in Stata

12 [46]. Regardless of the response variable, the statistically



significant predictors in the random-intercept models consistently

had large RI (chicken and dog blood index) in the multimodel

approach. To simplify the exposition, we report only the

multimodel analysis in the main text and show estimates based

on null-hypothesis testing in Table S3.

Results

Population abundanceOf 543 bugs from all stages collected at 94 houses with infested

human sleeping quarters, 17.7% were first- to third-instar nymphs;

12.7% fourth instars; 26.3% fifth instars; 21.2% females, and

22.1% males. Domestic bug abundance (median, 5 bugs per 0.5

person-hour) was highly variable (first-third quartiles, 2-9 bugs;

maximum, 41 bugs) at the 49 study houses and highly over-

dispersed (variance-to-mean ratio, 91.3/8.1 = 11.3). A median

study household had 5 people (first-third quartiles, 4–8), 2 dogs (2–

3), 0.5 cat (0–1), 15 chickens (7–22), 11 goats (0–29), 2 pigs (1–6)

and 2 horses (0–4); very young chickens were recorded in 71% of

houses. The proportion of the 49 study households that reported

allowing animals to sleep indoors was 55% (dogs), 37% (cats) and

35% (chickens). Chickens were allowed to roam freely, nesting

inside domiciles, kitchens, storerooms, chicken coops, or almost

anywhere around human sleeping quarters.

Bug abundance correlated positively and significantly with

numbers of dogs (r = 0.411, P,0.004) and cats (r = 0.291, P,

0.05), marginally with the total number of chickens (r = 0.280, P,

0.06), and non-significantly with human residents (r = 0.076, P.

0.6) or the indoor presence of chickens (r = 0.136, P.0.3).

Blood-feeding ratesOf 326 late-stage insects (from 49 houses) that were processed,

319 were examined for transparent urine. The percentage of

late-stage domestic T. infestans with transparent urine averaged

33.9% (108 of 319, 95% Confidence Interval [CI], 27.1–40.6%).

Temperature-adjusted feeding rates were nearly 4% lower than

unadjusted values (30.0%, CI, 23.6–36.6%). Daily blood-feeding

rates increased from 26.6% in fourth-instar nymphs to 36.2% and

34.6% in fifth instars and females, respectively, and declined to

21.7% in males (Figure 1A). Median blood-feeding intervals were

4.1 days, with large variations among households (first and third

quartiles, 2.4-7.0 days). A simple sensitivity analysis showed that a

10% perturbation of the parameters in the formula for the

correction factor c translated into close values for the daily feeding

intervals at 3.4 and 5.0 days (Text S1).

Daily blood-feeding rates decreased as the chicken blood index

increased (OR = 0.48, CI, 0.26-0.89), with marginal effects of male

stage (OR = 0.58, CI, 0.30–1.11)(Table 1, Figure 1B). There was

large model uncertainty (Table S4) and no empirical support for

effects due to mean maximum temperatures during the night

preceding bug capture (Figure S1), bug abundance, the dog blood

index and number of human residents (Table 1 and S1). The

averaged model fitted the data well (Hosmer and Lemeshow x2

test = 5.68, 8 df, P.0.6), and the area under the ROC was 0.615,

thus indicating a poor classification. None of the predictors had

significant regression coefficients when the chicken and dog blood

indices were replaced with the reported or observed presence of

chickens indoors and the household number of roommate dogs,

respectively (not shown).

Host-feeding choicesAll 319 T. infestans examined for transparent urine were

dissected and some blood meal residue extracted from 305 bugs

(from 47 houses). At least one host bloodmeal source was identified

in 289 (94.8%) of the bugs (from 45 houses); these 289 bugs were

termed ‘‘reactive’’ (bloodmeal identification data in Table S2).

Figure 1. Temperature-adjusted proportion of domestic T. infestans that blood-fed the night before catch according to bug stage(A) and the chicken blood index (B). Figueroa, October 2003 (spring). In B, each data point corresponds to a different house.doi:10.1371/journal.pntd.0002894.g001



The main bloodmeal sources were humans (68.2%, 95% CI, 53.4–

83.0%) followed by chickens (21.8%, 8.2–35.5%) and dogs (9.0%,

4.5–13.5%) (Figure 2). Blood meals on goats (2.1%, 0.4–4.5%),

pigs (1.4%, 0–2.8%), cats (1.0%, 0.5–2.6%) and rodents (0.3%, 0–

0.1%) were rare. Most reactive bugs had unmixed meals (95.2%),

a few had fed on two host species (4.5%) and hardly any on three

(0.3%). The proportion of bugs collected from human sleeping

quarters with unmixed blood meals from humans was very high

(65.4%, CI, 50.4–80.4%) compared with the series of 16 similar

studies compiled in [26]. The mean proportion of chicken-fed

domestic bugs clustered by site was five times greater in houses

where the indoor presence of chickens was reported or observed

(43.3%, CI, 18.7–68.0%) than in those where it was not (8.0%, CI,

0–18.9%). Among all the reactive bugs collected, the selection of

human versus chicken (or dog) bloodmeal sources was highly

significantly associated (Fisher’s exact tests, P,0.0001).

Human-feeding ratesThe proportion of human-fed bugs decreased strongly with an

increasing chicken blood index (OR = 0.010, CI, 0.003-0.033) and

with an increasing dog blood index (OR = 0.092, CI, 0.040–0.

211) (Table 1, Figure 3). There was no evidence that the number

of resident humans, bug stage, mean maximum temperatures

during the night preceding bug capture, and bug abundance per

site exerted any effects on the human blood index (Table 1 and

S1). The averaged model fitted the data very closely (Hosmer and

Lemeshow x2 test = 9.8, 8 df, P.0.28), and the area under the

ROC was 0.941 2an outstanding classification. Replacing the

chicken blood index with the presence of chickens indoors and the

dog blood index with roommate dog numbers yielded significant

regression coefficients for the indoor presence of chickens

(OR = 0.075, CI, 0.008–0.720) and marginally positive effects of

the number of human residents (OR = 12.72, CI, 0.86–188.99).

Hence, the indoor presence of chickens and dogs translated into a

lower human blood index.

The mean human-feeding rate per 100 bug-days was estimated

as 23.1 and the overall mean number of human-bug feeding

contacts per person-day as 7.2 (Table 2). When chickens were

available indoors, percent human-feeding rates were substantially

lower both in small and large (14.7–15.8) households than when

chickens were not reported or observed indoors (27.1–30.7)

(Kruskal-Wallis, df = 1, P,0.007), but there was large variation

within any group of households. The indoor presence of chickens

was also associated negatively with the frequency of human-

feeding contacts (Kruskal-Wallis, df = 1, P,0.001). Despite having

lower bug abundance, small households without chickens indoors

(42% of all infested houses) would experience two to five times

more human-bug feeding contacts per person-day (12.7) than

other households (2.7–5.2) in spring. One small household without

chickens indoors had an extreme value of 46.7 human-feeding

contacts per person-day. Excluding this extreme value did not alter

the qualitative relations between human-feeding rates or frequency

of feeding contacts with household size and indoor presence of

chickens.

The daily human-feeding rate also decreased dramatically with

increases in the chicken blood index (OR = 0.049, CI, 0.010–

0.236) and dog blood index (OR = 0.379, CI, 0.183–0.786)

(Figure 4, Table 1). Other predictors exerted no significant effects

(Table 1 and S1). The averaged model fitted the data very closely

(Hosmer and Lemeshow x2 test = 6.8, 8 df, P.0.5), and the area

under the ROC was 0.740 –an acceptable classification level. The

indoor presence of chickens was associated with a lower daily

human-feeding rate (OR = 0.414, CI, 0.191-0.900).

Discussion

Very high human blood indices and human-feeding rates of

domestic T. infestans decreased significantly with the increasing

frequency of blood meals on chickens and dogs or the occurrence

of chickens indoors in spring, in support of hypothesis 2 (i.e., the

human blood index and human-feeding rate should decrease with

more chicken or dog blood meals and the occurrence of indoor-

nesting chickens and dogs). As predicted by a mathematical model

of transmission, the data most strongly supported the hypothesis 4

that the rate of human-bug feeding contacts per person-day

decreased substantially with the indoor presence of chickens and

dogs [31].

These results –derived from combining information on daily

feeding rates and host blood indices in domestic areas with

household-level demographic data– support and extend previous

findings and predictions: high human blood indices in domestic T.

Table 1. Model-averaged coefficients of factors associated with the daily blood-feeding rate, human blood index, and dailyhuman-feeding rate of domestic T. infestans in Figueroa, spring 2003.

Daily blood-feeding ratea Human blood indexb Human-feeding ratec

Variablea Coefficient S.E. P RI Coefficient S.E. P RI Coefficient S.E. P RI

Intercept 20.8235 0.1750 *** 1.1690 0.2386 *** 21.6694 0.2623 ***

Chicken blood index 20.7294 0.3110 * 0.95 24.5767 0.5907 *** 1.00 23.0073 0.7973 *** 1.00

Dog blood index 20.3416 0.2863 ns 0.44 22.3817 0.4216 *** 1.00 20.9692 0.3713 ** 1.00

Stage: males 20.5625 0.3336 ms 0.45 0.2003 0.5135 ns 0.06 20.5178 0.3761 ns 0.31

Stage: females 0.0753 0.3157 ns 0.45 0.4139 0.5321 ns 0.06 0.1117 0.3456 ns 0.31

No. of humans 0.3704 0.2821 ns 0.39 20.0427 0.4664 ns 0.25 0.0206 0.2884 ns 0.19

Bug abundance 0.1743 0.2857 ns 0.26 20.6340 0.4815 ns 0.42 0.1910 0.2681 ns 0.28

Maximum temperature 20.1022 0.2884 ns 0.24 20.8591 0.5075 ns 0.60 20.0727 0.2892 ns 0.20

*** P,0.001; * P,0.05; ms, 0.05,P,0.1; ns, P.0.1.SE, standard error; RI, relative importance.aTotal number of observations with complete data for every predictor: 317 bugs from 47 houses.bTotal number of observations with complete data for every predictor: 289 bugs from 45 houses.cTotal number of observations with complete data for every predictor: 314 bugs from 44 houses.doi:10.1371/journal.pntd.0002894.t001



infestans were inversely related to chicken and dog blood indices in

surveys conducted elsewhere in Santiago del Estero during spring

and summer [2]. The large fraction of bugs with unmixed blood

meals implies that the bugs fed repeatedly on the same domestic

host species (host fidelity), which is consistent with previous

observations and the usual finding of bugs in the proximities of

host resting sites. Such increased human exposure is explained by

nightly resting patterns in winter and spring, when most local

villagers still slept indoors. In this region, chickens frequently are

allowed to nest indoors or are deliberately introduced there for

their protection; they have constant nesting locations during a few

weeks favoring bug feeding on them, and peak nesting frequency

in spring and early summer [47].

During the hot summer, however, people increasingly moved

their beds to sleep in the verandah or patio (which reduced their

exposure) and domestic bugs fed more often on dogs and chickens,

which reduced the human blood index. In the current study we

confirm that the same qualitative relationships hold in other rural

communities when domestic infestations were lighter and dogs

were less important bloodmeal sources than in earlier studies, and

extend those results to human-feeding rates.

Domestic bugs fed mostly on humans and chickens and had

fewer dog blood meals or mixed blood meals than in other studies

conducted in this region during late winter-early spring [26].

Differences in host exposure patterns and domestic animal

husbandry practices among settings may account for such

variations, given that bloodmeal identification methods (ELISA

versus agar double-diffusion tests) had similar performance in a

large-scale comparative trial of mosquito blood meals [48].

Compared with 16 published studies of domestic T. infestans, the

observed human blood index (68%) ranks tenth and equals the

mean human blood index found in the same study region in

Santiago del Estero over 1988-1992 [26]. The fraction of unmixed

blood meals in the current study (95%) is substantially higher than

the average across the 16 published studies (70%), and as a

consequence, the fraction of bugs with unmixed human blood

meals is comparatively very high (65.4%) [26].

Our study provides an improved way of estimating the daily

host-feeding rate of triatomine bugs and a measure of its precision

that are applicable to other settings and species of Triatominae.

The product of the human blood index (including both unmixed

and mixed blood meals) and daily blood-feeding rate is not

Figure 2. Percentage of late-stage T. infestans collected inhuman sleeping quarters that fed on each host species(regardless of feeding on other host species). Figueroa, spring2003. 95% confidence intervals clustered by site.doi:10.1371/journal.pntd.0002894.g002

Figure 3. Proportion of late-stage domestic T. infestans that fed on humans (the human blood index) according to the proportion ofbugs that fed on chickens (A) and dogs (B). Each data point corresponds to a different house. The human and chicken blood indices includebugs that fed on humans or chickens, respectively, regardless of feeding on other species. Figueroa, October 2003 (spring).doi:10.1371/journal.pntd.0002894.g003



dimensionally consistent; it is not a daily rate because the bugs

may have fed on humans (or on any other host species) over the

usually long period of time in which blood meals are detectable by

immunologic methods (i.e., up to three or four months, depending

on initial and subsequent bloodmeal sizes and temperature [49

and references therein]), and because the bugs may have fed once

or several times [26]. Therefore, the human-feeding rate

calculated on the basis of all human blood meals (mixed and

unmixed) is biased upward relative to the true rate because it may

include older human blood meals. The magnitude of the bias is

expected to increase with the proportion of mixed blood meals,

which in domestic T. infestans usually increases from a minimum in

late winter-early spring (as in the current study) to late summer,

and is quite high throughout the species’ range [26,50].

Conversely, the unmixed human blood index among bugs that

fed on the preceding night is a more accurate estimate of the daily

human-feeding rate. To reduce the potential bias and detect

mixed blood meals, at least all resident host species in the study

habitats need to be included in sensitive bloodmeal identification

tests. These considerations also apply to the derived human-

feeding rate per person.

Our estimate of the average blood-feeding interval of domestic

T. infestans in spring (4.1 days) is higher than another spring-

summer average (3 days) [35], and considerably lower than

another estimate for domestic adult bugs (5–8 days) in two heavily

infested houses [33]. This high feeding frequency is consistent with

experimental work showing that when a live rabbit is continually

present T. infestans and R. prolixus females will feed about every

three days [51]. Moreover, there were large variations in blood-

feeding rates among houses. The top models for blood-feeding

rates had considerable model uncertainty (i.e., low Akaike weights)

and less support than the models for the human blood index and

human-feeding rate. According to the averaged model, the more

the bugs fed on chickens the less frequently they fed. This result

Table 2. Human-feeding rate per 100 bug-days and number of human-bug feeding contacts per person-day of late-stagedomestic T. infestans according to presence of chickens indoors and numbers of resident people (Figueroa, spring 2003).

No. ofpeople

Presence ofchickens indoors

No. ofhousesa

Mean bug abundanceper person-hour (SE)

Human-feeding rateper 100 bug-days (CI)

No. of human-bug feeding contactsper person-day (CI)

1–5 Yes 7 8.7 (2.6) 14.7 (0.2–29.2) 4.0 (0.2–7.9)

1–5 No 20 7.3 (2.1) 30.7 (21.1–40.2) 12.7 (0–30.3)

6–17 Yes 10 10.6 (4.1) 15.8 (0.1–43.3) 2.7 (0–6.6)

6–17 No 11 7.5 (2.6) 27.1 (10.9–43.3) 5.2 (2.6–7.7)

Total 48 8.1 (1.4) 23.1 (15.7–30.5) 7.2 (0.3–14.2)

aExcludes one house with no human occupants.CI, 95% confidence interval.doi:10.1371/journal.pntd.0002894.t002

Figure 4. Daily human-feeding rate of late-stage domestic T. infestans according to the proportion of chicken-fed bugs (A) and dog-fed bugs (B). The daily human-feeding rate is the proportion of bugs that fed on humans only on the previous night. Figueroa, October 2003(spring). Each data point corresponds to a different house.doi:10.1371/journal.pntd.0002894.g004



was surprising and might be related to an eventually larger

bloodmeal size of bugs feeding on chickens; if bugs achieved a

higher engorgement status when feeding on chickens, then they

would not need to feed as often as bugs that ingested smaller blood

meals. In the absence of direct measures of bloodmeal size, this

explanation is tentative and the subject needs to be more widely

investigated. Male bugs tended to feed less often than females [52]

or late nymphal instars which require more energy for laying eggs

and molting. Unlike longer experimental studies [34,38], no

temperature or host abundance effects on daily feeding rates were

detected despite considerable variations in temperature among

catch days (range, 13uC) and in domestic host abundance among

households (range, 2–17), thus rejecting hypothesis 1. Lack of

effects of bug abundance on blood-feeding rates, human blood

indices and human-feeding rates (hypothesis 3) may be explained

by the rather limited range of variation in domestic bug

abundance at the beginning of the reproductive season a few

years after vector control actions, and the imprecision of bug

abundance estimates by timed manual collections. The evidence

collectively indicates that the high blood-feeding rate of domestic

T. infestans is little affected by temperature, stage, numbers of

domestic hosts and domestic bug abundance in this specific

context in the dry Argentine Chaco spring.

The estimated number of human-bug feeding contacts per

person-day averaged 7.2 (median, 3.6) over households and was

quite high. If the bugs bite only at night, this means nearly one

human-bug feeding contact per person every one to two hours

during the night, but more extreme values were recorded. These

estimates do not include first- to third-instar nymphs, which

comprise an increasingly larger fraction of the bug population

from spring to summer. First- to third-instar nymphs are easily

captured in human beds, and most are fed on humans [26,28,41].

These small instars may thus increase substantially the total

human-bug contact rate, ensuing blood loss, host irritation, and

inoculation of bug saliva with potentially allergic effects [53].

Uncertainty about the capture efficiency of bugs per unit of effort

limits the absolute value of estimates for each of these quantities,

but still allows a direct comparison among households. For

example, the proportion of existing late-stage domestic R. prolixus

collected manually without a dislodgant spray varied from 4 to

41% depending on bug stage and type of walls and roof, and was

invariant with absolute bug abundance [54]. The estimated daily

number of human-bug feeding contacts also puts a cap on the

frequency of potentially infective contacts experienced by an

average person because late–stage bugs also concentrate most

infections with T. cruzi [25,55].

Our study has both limitations and strengths. The estimated

blood-feeding intervals may be taken as good approximations to

the actual values. In addition to the pioneering studies of Catala

[34] using live pigeons, our subsequent experiments using

heparinized cow blood and an artificial feeding apparatus showed

that transparent urine was detected regardless of bloodmeal size in

.95%, 20% and none of a total of 729 fourth- and fifth-instars

nymphs of T. infestans held at 26–28uC and examined at 12, 36 and

60 h post-feeding, respectively (P. L. Marcet et al., unpublished

results). These results are in close agreement with the predicted

values at 25uC [34] 2the average overnight temperature during

the current field study2 despite major differences in host species,

host blood composition (i.e., bird versus mammal) and experi-

mental setup. Our experiments also showed that transparent urine

was detectable at 12 h post-feeding in 90% of adult females that

had achieved full blood meals, and then persisted only in 4% of

them at 36 h post-feeding (total examined, 285 females); partial

blood meals slightly reduced the rate of occurrence of transparent

urine after feeding. The achieved bloodmeal sizes of adult bugs

under field conditions are unknown and may be used to improve

estimates of blood-feeding. This method would not be able to

reveal the occurrence of multiple blood meals during the same

night. T. cruzi infection does not modify the rate of urine excretion

in triatomine bugs, and affects minimally their vital rates only

under severe starvation conditions [56,57], which was not the case

for the study bug populations. The experimental effects of T. cruzi

infection on blood-feeding rates and blood intake of T. infestans

were minor, temperature-dependent and confined to fifth instars

[56,58]. Although the correction factor c was calibrated in the

range between 18 and 28uC whereas overnight average maximum

temperatures sometimes exceeded 30uC, the linear relationship

between c and temperature has strong underlying physiological

foundations: The rate of urine excretion (diuresis) in R. prolixus was

constant and linearly related to temperature within the range 17–

30uC, and responded almost immediately to temperature changes

[59].

Although molecular methods represent an improvement over

ELISA for bloodmeal identification at host genus or species levels,

different PCR protocols frequently fail to detect host bloodmeal

sources in a sizable (15–60%) proportion of tested bugs

[e.g.,50,60,61] and older blood meals. This is likely related to

degradation of host DNA during the blood meal’s long periods of

storage in the anterior midgut. Unlike serologic methods for

bloodmeal identification [49], genus- or species-specific PCRs used

to detect the persistence of host DNA in T. infestans experimentally

fed on major domestic host species identified host sources

consistently only up to two weeks post-feeding, and to a much

lesser extent up to four weeks post-feeding [62]. For domestic bug

populations exposed to a limited, well-known range of domestic

host species, the ELISA test is a cost-effective option showing high

sensitivity and specificity combined with high-throughput process-

ing. It revealed bloodmeal residues that were visually undetectable

and a sizable fraction of mixed blood meals on dogs and chickens

or cats housed overnight in pairs in experimental huts [40]. Here

the ELISA test kept the proportion of non-reactive bugs below

5%, yet it may have missed marginal blood meals on other

infrequent hosts (e.g., horses). Future studies may benefit from

collecting larger samples of bugs per site to increase the precision

of estimates.

Host occupancy changes over time and is difficult to gauge

precisely [37], especially in peridomestic sites, which limits our

understanding of host-feeding choices and related metrics. The

question of which factors are causing host shifts or the lack of them

(e.g., host availability and accessibility, bug attack rates, temper-

ature, refuge availability and its interactions) remains unresolved.

Major strengths of this study are the detailed information on

various bug attributes in the identified sleeping quarters of a

sizable number of houses from a well-defined rural area, and

appropriate GLMM-based modeling of clustered data in a

multimodel inferential framework.

Implications for vector control and disease transmissionWe have not assessed the links between host-feeding rate or host

choice and household infection with T. cruzi, a central goal that

requires a much greater research effort. However, some evidence

allows us to make inferences about parasite transmission. Our

study shows a very intense human-bug contact rate in spring that is

consistent with the peak of monthly acute cases of Chagas disease

recorded historically in October and November in northern

Argentina [63,64]. Other field and experimental data also showed

that the abundance of domestic T. infestans, its prevalence of

infection with T. cruzi, and the mean intensity of parasites in bugs’



rectal contents increased very fast from a minimum at the end of

winter to high or submaximal values by the end of spring

[25,64,65]. Average temperatures during our study are considered

optimal for T. infestans. Because domestic bugs feed dispropor-

tionately more on humans at the beginning of the reproductive

season in late winter and spring than during the hot summer

season when bug density peaks [31], spring is a high-risk period for

increased domestic transmission of T. cruzi.

Although a snapshot of the transmission system indicates that

indoor chickens and dogs in the spring lower (at least temporarily)

the probability of vectors feeding on transmission-competent hosts

and human-bug feeding contacts (supporting hypotheses 2 and 4),

it fails to capture the entire range of dynamic effects on parasite

transmission rates over time. The indoor presence of dogs in

spring may increase human risks of infection because the

infectivity of T. cruzi-infected dogs is 18.7 times greater than that

of infected humans [21]. The increased spring bug population fed

on indoor-resting chickens, humans and dogs [28] would increase

the human-bug contact rate during the subsequent summer when

the nesting activity of chickens declines and chickens move away

from human sleeping quarters [25,31] while most people move

outside to sleep in their verandahs. Based on this chain of effects,

each individually well documented, we hypothesize (as confirmed

by 1992 summer data [25,29] from Amama, Santiago del Estero,

Argentina, and in accordance with our mathematical model of

transmission [31]) that the net effects of chickens on parasite

transmission in the presence of transmission-competent hosts may

be more adequately described by zoopotentiation [13] than by

zooprophylaxis. This hypothesis cannot be regarded as tested in

Figueroa until quantitative data from Figueroa on the prevalence

and transmission of infection in bugs, humans, and other hosts

during the summer months, or over an annual cycle, are analyzed.

The large variability among households in blood-feeding rates

and human blood indices implies large uncertainty in the

threshold human-feeding rate of domestic T. infestans under which

no transmission would occur. Measurement of a threshold human-

feeding rate for transmission of vector-borne pathogens is fraught

with several sources of inaccuracy, including measurement of

vector abundance, host exposure and the probability of human

infection given a feeding contact with an infected vector, for which

only indirect estimates are possible [5,24,31,41,66]. Estimates of

transmission thresholds using house infestation prevalence at a

village-wide level [67] are even more removed from the basic

quantities and processes that determine the theoretical thresholds

and should be regarded with reservations until solid evidence is

available.

The presence of domestic animals in human sleeping quarters

and changing site occupancy by different domestic host species

exert profound effects on the population dynamics of T. infestans,

host-feeding choices and ensuing risks of parasite transmission.

The evidence available thus far advises against the presence of

domestic animals in human sleeping quarters [24,25,31]. Treating

chickens and dogs with insecticides that have reduced repellency

may turn them into baited lethal traps and exert substantial

impacts on infestation and transmission [68]. Appropriate animal

husbandry combined with long-lasting environmental modifica-

tions in housing and peridomestic structures [69] and judicious use

of residual insecticide spraying are crucial for sustainable vector

and disease control of Chagas disease in affected rural areas [21].

The human-feeding rates of major domestic vectors of T. cruzi

should be estimated more widely with currently available methods.

Whether zooprophylaxis or zoopotentiation applies to other

domestic triatomine species in other regions and ecoepidemiolo-

gical circumstances merits further research.

Supporting Information

Figure S1 Temperature-adjusted proportion of domes-tic T. infestans that blood-fed the night before catch(daily feeding rate) according to mean maximumtemperature during that night. Figueroa, October 2003

(spring).

(TIF)

Figure S2 Human blood index (A) or human-feedingrate (B) according to daily feeding rates of domestic T.infestans for all houses. Figueroa, October 2003 (spring).

(TIF)

Table S1 Weather data during bug collection surveys.Figueroa, October 2003 (spring).

(XLS)

Table S2 Bloodmeal identification results of domesticT. infestans as determined by direct ELISA tests.Figueroa, October 2003 (spring). Codes 0 and 1 are for negative

and positive identification results, respectively; ND, no data.

(XLS)

Table S3 Random-intercept multiple logistic regressionobtained with Stata.

(DOC)

Table S4 Multimodel assessment of factors associatedwith daily blood-feeding rates, human blood index, andhuman-feeding rates of T. infestans. Figueroa, spring 2003.

(DOC)

Text S1 Sensitivity analysis of blood-feeding rates tochanges in parameter estimates of the correction factor.

(DOC)

Acknowledgments

In memory of Joaquın Zarate, head of the National Vector Control

Program at Tucuman. We thank Leonardo Lanati and Natalia

Macchiaverna for providing active support; Padre Sergio, Sara and Tuki

family of Bandera Bajada for field accommodation, Silvana Ferreyra for

technical assistance to set up the database. Ricardo E. Gurtler and Marıa

C. Cecere are members of the CONICET Researcher’s Career. JEC

acknowledges with thanks the assistance of Priscilla K. Rogerson, and the

hospitality of the family of William T. Golden during this work.

Author Contributions

Conceived and designed the experiments: REG MCC LAC GMVP JEC.

Performed the experiments: MCC GMVP LAC JMG MdPF. Analyzed the

data: REG MdPF GMVP. Contributed reagents/materials/analysis tools:

REG MCC GMVP MdPF UK JEC. Wrote the paper: REG UK JEC.

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Domestic animal hosts strongly influence human-feeding rates of the chagas disease vector Triatoma...

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